Javier Sancho

University of Zaragoza | UNIZAR · Department of Biochemistry and Molecular and Cellular Biology

Protein design requires a deep control of protein folding energetics, which can be determined experimentally on a case-by-case basis but is not understood in sufficient detail. Calorimetry, protein engineering and biophysical modeling have outlined the fundamentals of protein stability, but these approaches face difficulties in elucidating the spec...

Despite impressive advances by AlphaFold2 in the field of computational biology, the protein folding problem remains an enigma to be solved. The continuous development of algorithms and methods to explore longer simulation timescales of biological systems, as well as the enhanced accuracy of potential functions (force fields and solvent models) hav...

In molecular dynamics simulations we can often increase the time step by imposing constraints on internal degrees of freedom, such as bond lengths and bond angles. This allows us to extend the length of the time interval and therefore the range of physical phenomena that we can afford to simulate. In this article we analyse the impact of the accura...

Molten globule (MG) is the name given to a compact, non‐native conformation of proteins that has stimulated the imagination and work in the protein folding field for more than 40 years. The MG has been proposed to play a central role in the folding reaction and in important cell functions, and to be related to the onset of misfolding diseases. Due...

Protein stability is a requisite for most biotechnological and medical applications of proteins. As natural proteins tend to suffer from a low conformational stability ex vivo, great efforts have been devoted toward increasing their stability through rational design and engineering of appropriate mutations. Unfortunately, even the best currently us...

Phenylketonuria (PKU) is a rare metabolic disease caused by variations in a human gene, PAH, encoding phenylalanine hydroxylase (PAH), and the enzyme converting the essential amino acid phenylalanine into tyrosine. Many PKU-causing variations compromise the conformational stability of the encoded enzyme, decreasing or abolishing its catalytic activ...

PirePred is a genetic interpretation tool for a variety of medical conditions investigated in newborn screening programs. The PirePred server retrieves, analyses and displays in real time genetic and structural data on 58 genes/proteins associated with medical conditions frequently investigated in the newborn. PirePred analyses the predictions gene...

The treatment of Parkinson’s disease (PD), the second most common neurodegenerative human disorder, continues to be symptomatic. Development of drugs able to stop or at least slowdown PD progression would benefit several million people worldwide. SynuClean-D is a low molecular weight 2-pyridone-based promising drug candidate that inhibits the aggre...

Antimicrobial resistant (AMR) bacteria constitute a global health concern. Helicobacter pylori is a Gram-negative bacterium that infects about half of the human population and is a major cause of peptic ulcer disease and gastric cancer. Increasing resistance to triple and quadruple H. pylori eradication therapies poses great challenges and urges th...

Human phenylalanine hydroxylase (PAH) is a metabolic enzyme involved in the catabolism of L-Phe in liver. Loss of conformational stability and decreased enzymatic activity in PAH variants result in the autosomal recessive disorder phenylketonuria (PKU), characterized by developmental and psychological problems if not treated early. One current ther...

Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiac disease. Variants in MYBPC3, the gene encoding cardiac myosin-binding protein C (cMyBP-C), are the leading cause of HCM. However, the pathogenicity status of hundreds of MYBPC3 variants found in patients remains unknown, as a consequence of our incomplete understanding of the pa...

Background Human metapneumovirus (HMPV) is an important etiologic agent of respiratory tract infection (RTI). This study aimed to describe its genetic diversity and clinical impact in patients attended at a tertiary university hospital in Barcelona from the 2014-2015 to the 2016-2017 seasons, focusing on the emerging duplications in G gene and thei...

Context Only two mutations at the Lysine 183 amino acid in the extracellular N‐terminal domain of human TSH receptor (hTSHR) have been associated to hypersensitivity to hCG and familial gestational hyperthyroidism. Patients A 38‐year‐old woman was seen during the first trimester of her second pregnancy for thyrotoxicosis with increased fT3 and fT4...

Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiac disease. Mutations in MYBPC3, the gene encoding cardiac myosin-binding protein C (cMyBP-C), are a leading cause of HCM. However, it remains challenging to define whether specific gene variants found in patients are pathogenic or not, limiting the reach of cardiovascular genetics...

The invention relates to 4-phenyl dihydropyridine deriva­tives of formula I, and to pharmaceutical compositions of same, wherein the meaning of R1, R2, R3 and A is that indicated in the description, for the treatment and/or prevention of an infection or disease caused by said infec­tion, the infection being caused by the bacteria Helicobacter.

Helicobacter pylori is considered the most prevalent bacterial pathogen in humans. The increasing antibiotic resistance evolved by this microorganism has raised alarm bells worldwide due to the significant reduction in the eradication rates of traditional standard therapies. A major challenge in this antibiotic resistance crisis is the identificati...

Flavodoxins are small soluble electron transfer proteins widely present in bacteria and absent in vertebrates. Flavodoxins participate in different metabolic pathways and, in some bacteria, they have been shown to be essential proteins representing promising therapeutic targets to fight bacterial infections. Using purified flavodoxin and chemical l...

Antibiotic resistance is a major cause of the increasing failures in the current eradication therapies against Helicobacter pylori. In this scenario, repurposing drugs could be a valuable strategy to fast-track novel antimicrobial agents. In the present study, we analyzed the inhibitory capability of 1,4-dihydropyridine (DHP) antihypertensive drugs...

The increasing ease with which massive genetic information can be obtained from patients or healthy individuals has stimulated the development of interpretive bioinformatics tools as aids in clinical practice. Most such tools analyze evolutionary information and simple physical-chemical properties to predict whether replacement of one amino acid re...

α-Synuclein (α-Syn) forms toxic intracellular protein inclusions and transmissible amyloid structures in Parkinson’s disease (PD). Preventing α-Syn self-assembly has become one of the most promising approaches in the search for disease-modifying treatments for this neurodegenerative disorder. Here, we describe the capacity of a small molecule (ZPD-...

As proteins perform most cellular functions, quantitative understanding of protein energetics is required to gain control of biological phenomena. Accurate models of native proteins can be obtained experimentally but the lack of equally fine models of unfolded ensembles impedes the calculation of protein folding energetics from first principles. He...

The increasing antibiotic resistance evolved by Helicobacter pylori has alarmingly reduced the eradication rates of first-line therapies. To overcome the current circulating resistome, we selected a novel potential therapeutic target in order to identify new candidate drugs for treating H. pylori infection. We screened 1120 FDA-approved drugs for m...

Helicobacter pylori ( Hp) infection is the main cause of peptic ulcer and gastric cancer. Hp eradication rates have fallen due to increasing bacterial resistance to currently used broad-spectrum antimicrobials. We have designed, synthesized, and tested redox variants of nitroethylene- and 7-nitrobenzoxadiazole-based inhibitors of the essential Hp p...

Aims: The human apoptosis-inducing factor (hAIF) supports OXPHOS biogenesis and programmed cell death, with missense mutations producing neurodegenerative phenotypes. hAIF senses the redox environment of cellular compartments, stabilizing a charge transfer complex (CTC) dimer that modulates the protein interaction network. In this context, we aimed...

GSK3082 – a hepatitis C virus RNA polymerase inhibitor – and a series of analogues with structural diversity at the 5-position were prepared from a 2,2,4,5-tetrasubstituted pyrrolidine obtained with a well-defined stereochemistry from the 1,3-dipolar cycloaddition of the chiral imino ester derived from leucine tert-butyl ester and (R)-2,3-O-isoprop...

Amyloid fibrils formed by a variety of peptides are biological markers of different human diseases, such as Alzheimer’s disease, Parkinson’s disease, and type II diabetes, and are structural constituents of bacterial biofilms. Novel fluorescent probes offering improved sensitivity or specificity toward that diversity of amyloid fibrils or providing...

Significance Parkinson’s disease is characterized by the accumulation of amyloid deposits in dopaminergic neurons, mainly composed of the protein α-synuclein. The disordered nature of α-synuclein and its complex aggregation reaction complicate the identification of molecules able to prevent or revert the formation of these inclusions and the subseq...

Ferredoxin-NADP(H) reductases (FNRs) deliver NADPH or low potential one-electron donors to redox-based metabolism in plastids and bacteria. Xanthomonas citri subsp. citri (Xcc) is a Gram-negative bacterium responsible for citrus canker disease that affects commercial citrus crops worldwide. The Xcc fpr gene encodes a bacterial type FNR (XccFPR) tha...

Near the minimum free energy basin of proteins where the native ensemble resides, partly unfolded conformations of slightly higher energy can be significantly populated under native conditions. It has been speculated that they play roles in molecular recognition and catalysis, but they might represent contemporary features of the evolutionary proce...

An increasing number of neurodegenerative diseases are being found to be associated with the abnormal accumulation of aggregated proteins in the brain. In Parkinson’s disease, this process involves the aggregation of alpha-synuclein (α-syn) into intraneuronal inclusions. Thus, compounds that inhibit α-syn aggregation represent a promising therapeut...

Streptococcus pneumoniae (Sp) strain TIGR4 is a virulent, encapsulated serotype that causes bacteremia, otitis media, meningitis and pneumonia. Increased bacterial resistance and limited efficacy of the available vaccine to some serotypes complicate the treatment of diseases associated to this microorganism. Flavodoxins are bacterial proteins invol...

There exist two isoforms of cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C) in pig populations that differ in a single amino acid (Met139Leu). The isoenzymes have different kinetic properties, affecting more strongly the Km and Vmax of nucleotides. They are associated to different phenotypes modifying traits of considerable economic interest....

Dixon plots of 3-MP adding PEP and GDP as variable substrates. (A) Dixon plot of 139Met varying PEP. (B) Dixon plot of 139Leu varying PEP. (C) Dixon plot of 139Met at saturating concentration of GDP. (D) Dixon plot of 139Leu at saturating concentration of GDP. (TIF)

Concentration-dependent inhibition of PEPCK-C by 3-MP. Data are the mean of three independent experiments ± SD. (TIF)

Concentration-dependent inhibition of PEPCK-C by compounds 1 and 2. Data are the mean of three independent experiments ± SD. Compound 1 plot (A) was adjusted to linear regression due to its precipitation above 200 μM. (TIF)

Dixon plots of 3-MP adding OAA and GTP as variable substrates. (A) Dixon plot of 139Met varying OAA. (B) Dixon plot of 139Leu varying OAA. (C) Dixon plot of 139Met at saturating concentration of GTP. (D) Dixon plot of 139Leu at saturating concentration of GTP. (TIF)

Human Amylin, or islet amyloid polypeptide (hIAPP), is a small hormone secreted by pancreatic β-cells that forms aggregates under insulin deficiency metabolic conditions, and it constitutes a pathological hallmark of type II diabetes mellitus. In type II diabetes patients, amylin is abnormally increased, self-assembled into amyloid aggregates, and...

Experimental and computational screenings are currently widespread tools for identifying either potential ligands for a given target or potential targets for a given chemical compound. In particular, ligand-induced stabilization against thermal denaturation (or thermal shift assay) is an easy and convenient experimental procedure for finding compou...

Familial Hypercholesterolemia (FH), a genetic disorder with a prevalence of 0.2 %, represents a high risk factor to develop cardiovascular and cerebrovascular diseases. The majority and most severe FH cases are associated to mutations in the receptor for Low Density Lipoproteins (LDL-r), but the molecular basis explaining the connection between mut...

In many cases the stability of a protein has to be increased to permit its biotechnological use. Rational methods of protein stabilization based on optimizing electrostatic interactions have provided some fine successful predictions. However, the precise calculation of stabilization energies remains challenging, one reason being that the electrosta...

The LDL receptor (LDLR) internalizes LDL and VLDL particles In the endosomes, it adopts a closed conformation important for recycling, by interaction of two modules of the Ligand Binding domain (LR4-5) and a β-propeller motif. Here, we investigate by SPR the interactions between those two modules and the β-propeller. Our results indicate that the t...

Binding/unbinding of small ligands, such as ions, to/from proteins influences biochemical processes such as protein folding, enzyme catalysis or protein/ligand recognition. We have investigated the mechanism of chloride/water exchange at a protein surface (that of the apoflavodoxin from Helicobacter pylori) using classical all-atom molecular dynami...

Aβ is intensely studied to characterize its conformations and find drugs to block its key roles in Alzheimer´s disease. While Aβ should be unfolded at the start of such studies, achieving this state is difficult as the peptide tends to adopt aggregating β-structures. Analysis of Aβ1-40´s 13C, 15N and 1Hα chemical shifts and 1HN-1HN NOEs in DMSO, re...

The LDL receptor internalizes circulating LDL and VLDL particles for degradation. Its extracellular binding domain contains ten (seven LA and three EGF) cysteine-rich modules, each bearing three disulfide bonds. Despite the enormous number of disulfide combinations possible, LDLR oxidative folding leads to a single native species with 30 unique int...

The cytochrome b559 is a heme-bridged heterodimeric protein with two subunits, α and β. Both subunits from Synechocystis sp. PCC 6803 have previously been cloned and overexpressed in Escherichia coli and in vivo reconstitution experiments have been carried out. The formation of homodimers in the bacterial membrane with endogenous heme was only obse...

Osmolytes are cosolutes that induce stabilization of the structure of proteins. Polyols are osmolytes that have importance in a wide variety of biotechnological and industrial processes. In this work, a systematic study concerning the effect of polyols of different number of methylene and hydroxyl groups on the stability of α-chymotrypsinogen A is...

Increasing the thermostability of proteins is often crucial for their successful use as analytic, synthetic or therapeutic tools. Most rational thermostabilization strategies were developed on small two-state proteins and, unsurprisingly, they tend to fail when applied to the much more abundant, larger, non-fully cooperative proteins. We show that...

The Low Density Lipoprotein receptor (LDLR) captures circulating lipoproteins and delivers them in the endosome for degradation. Its function is essential for cholesterol homeostasis, and mutations in the LDLR are the major cause of Familiar Hypercholesterolemia. The release of LDL is usually attributed to endosome acidification. As the pH drops, t...

In the filamentous cyanobacterium Anabaena sp. PCC 7120, the ferric uptake regulator FurA functions as a global transcriptional regulator. Despite several analyses have focused on elucidating the FurA-regulatory network, the number of target genes described for this essential transcription factor is limited to a handful of examples. In this article...

Background: Prions are a particular type of amyloids related to a large variety of important processes in cells, but also responsible for serious diseases in mammals and humans. The number of experimentally characterized prions is still low and corresponds to a handful of examples in microorganisms and mammals. Prion aggregation is mediated by spe...

The molecular mechanism of lipoprotein binding by the LDL receptor (LDLR) is poorly understood, one reason being structures of lipoprotein-receptor complexes are not available. LDLR uses calcium binding repeats (LR) to interact with apolipoprotein B and apolipoprotein E (apo B and apo E). We have characterized by NMR and SPR the complexes formed by...

Helicobacter pylori is a gram-negative bacterium that infects the luminal surface of the human gastric epithelium. Around one half of the world’s population is thought to be infected by this bacterium, which is able to develop diseases such as peptic ulcer or gastric cancer. Eradication of Helicobacter pylori is becoming increasingly difficult due...

The nonstructural protein 3 (NS3) from the hepatitis C virus processes the non-structural region of the viral precursor polyprotein in infected hepatic cells. The NS3 protease activity has been considered a target for drug development since its identification two decades ago. Although specific inhibitors have been approved for clinical therapy very...

Helicobacter pylori (Hp) infection affects one half of the human population and produces a variety of diseases from peptic ulcer to cancer. Current eradication therapies achieve modest success rates (around 70 %), resistances to the antibiotics of choice are on the rise, and vaccination has not proved successful yet. Using an essential Hp protein,...

Background Prion proteins conform a special class among amyloids due to their ability to transmit aggregative folds. Prions are known to act as infectious agents in neurodegenerative diseases in animals, or as key elements in transcription and translation processes in yeast. It has been suggested that prions contain specific sequential domains with...

Helicobacter pylori is a globally important and genetically diverse gastric pathogen that infects most people in developing countries. Eradication efforts are complicated by antibiotic resistance, which varies in frequency geographically. There are very few data on resistance in African strains. Sixty-four Gambian H. pylori strains were tested for...

Alzheimer's disease, characterized by deposits of amyloid β-peptide (Aβ), is the most common neurodegenerative disease, but it still lacks a specific treatment. We have discovered five chemically unrelated inhibitors of the in vitro aggregation of the Aβ17−40 peptide by screening two commercial chemical libraries. Four of them (1−4) exhibit relativ...

Protein stability is not just an academic matter. Biotechnology, Cell Biology and Drug Design are some of the fields where both theoretical and practical knowledge of protein stability is required. Simple equipment and chemicals, such as a thermostated fluorimeter and common denaturants, suffice to determine the conformational stability of a protei...